Resistance to normal regulated cell death (apoptosis) is one of the characteristics of the cancer phenotype. Escape from apoptosis allows cancer cells to grow under conditions of low oxygen, high acidity and limited nutrient supply such as are found in the growing tumor where normal cells cannot survive. It also explains the intrinsic resistance of cancer cells to radiation and chemotherapy. Understanding the mechanisms by which cancer cells become resistant to apoptosis is central to the development of new therapies for the treatment of cancer and for overcoming drug resistance. Thioredoxins (Trxs) are small redox proteins that undergoes reversible NADPH- dependent reduction by flavoprotein thioredoxin reductases. There are two human Trxs, a cytosolic Trx-1 and a mitochondrial Trx-2. Trx-1 transforms normal cells, stimulates cell growth and causes a marked resistance to apoptosis by a variety of anticancer drugs. Trx-1 is over expressed in many human primary tumors where it is associated with aggressive tumor growth, decreased spontaneous apoptosis and with decreased patient survival. The mechanism(s) by which Trx-I inhibits apoptosis is not known. We have shown that Trx-1 binds to and represses the activity of the tumor suppresser protein PTEN, a lipid phosphatase inhibitor of the phosphatidyinositol-3-kinase/Akt cell survival signaling pathway. Trx-1 also decreases the expression of Noxa a regulator ofp53-dependent apoptosis. We present new evidence that Trx-2 protects mitochondria against oxidant damage and may protect cancer cells against mitochondrial mediated apoptosis. The hypothesis upon which the proposed studies are based is that the Trxs play a critical role in preventing apoptosis in cancer cells; Trx-1 by inactivation of the phosphatidyinositol-3-kinase/Akt cell survival pathway and p53-mediated apoptosis; and Trx-2 by protection against mitochondrial damage. Furthermore, we propose that inhibitors of Trx will block these antiapoptotic effects and will be effective antitumor agents. We will use the Trx-1 inhibitor PX-12 and two novel Trx-1 inhibitors we have identified. The objective of our studies is to investigate mechanisms for Trx inhibition of apoptosis and develop novel inhibitors of Trx we have identified as potential agents for the treatment of cancer